This proposal describes an outstanding research program designed to continue electron microscopy structural studies on a number of important nucleo-protein complexes involved in the process of genetic recombination in both prokaryotic and eukaryotic cells. The work proposed here will be focused on the study of 1) filamentous complexes such as E. coli RecA-DNA complexes and its eukaryotic analog, Rad 51, which is likely to play a key role in recombination in yeast and also in humans; and 2) hexameric helicases, proteins that catalyze the opening of the DNA double strand to form single strand, utilizing the energy of ATP hydrolysis. Some of the proteins of this family have been characterized in Dr. Egelman's laboratory in the previous funding period. In both these cases, Dr. Egelman proposes to: 1) Improve the spatial resolution of the three-dimensional reconstruction of these nucleoprotein complexes, as derived from conventional, cryo and 3-D reconstruction electron microscopy methods. For the filaments, Dr. Egelman proposes to improve the resolution of the reconstructions by complexing the helical assembly with other proteins (such as UmuD, antibodies, Rad52, etc.) and also by examining other assemblies such as Dmc1. Improved resolution of the filament will be necessary to characterize the conformational change involved in the transition from inactive (for which X-ray data exist) to active RecA filaments (for which no crystal has been obtained). 2) To characterize the interactions between the helical filaments and the hexameric proteins with other proteins, in particular: complexes of RecA with UmuD, DnaB with DnaG, Rad51 with Rad52. 3) Scan for homologous proteins in other organisms to test the hypothesis, validated in part by the results obtained during the previous funding period, that both the helical filaments and the hexameric ring structure of the helicases are structures highly conserved throughout evolution.
| López-Castilla, Aracelys; Thomassin, Jenny-Lee; Bardiaux, Benjamin et al. (2017) Structure of the calcium-dependent type 2 secretion pseudopilus. Nat Microbiol 2:1686-1695 |
| Wang, Fengbin; Burrage, Andrew M; Postel, Sandra et al. (2017) A structural model of flagellar filament switching across multiple bacterial species. Nat Commun 8:960 |
| Zheng, Weili; Wang, Fengbin; Taylor, Nicholas M I et al. (2017) Refined Cryo-EM Structure of the T4 Tail Tube: Exploring the Lowest Dose Limit. Structure 25:1436-1441.e2 |
| Kasson, Peter; DiMaio, Frank; Yu, Xiong et al. (2017) Model for a novel membrane envelope in a filamentous hyperthermophilic virus. Elife 6: |
| Frenz, Brandon; Walls, Alexandra C; Egelman, Edward H et al. (2017) RosettaES: a sampling strategy enabling automated interpretation of difficult cryo-EM maps. Nat Methods 14:797-800 |
| Subramaniam, Sriram; Earl, Lesley A; Falconieri, Veronica et al. (2016) Resolution advances in cryo-EM enable application to drug discovery. Curr Opin Struct Biol 41:194-202 |
| Costa, Tiago R D; Ilangovan, Aravindan; Ukleja, Marta et al. (2016) Structure of the Bacterial Sex F Pilus Reveals an Assembly of a Stoichiometric Protein-Phospholipid Complex. Cell 166:1436-1444.e10 |
| Lu, Alvin; Li, Yang; Yin, Qian et al. (2015) Plasticity in PYD assembly revealed by cryo-EM structure of the PYD filament of AIM2. Cell Discov 1: |
| DiMaio, Frank; Chen, Chun-Chieh; Yu, Xiong et al. (2015) The molecular basis for flexibility in the flexible filamentous plant viruses. Nat Struct Mol Biol 22:642-4 |
| DiMaio, Frank; Yu, Xiong; Rensen, Elena et al. (2015) Virology. A virus that infects a hyperthermophile encapsidates A-form DNA. Science 348:914-7 |
Showing the most recent 10 out of 71 publications
